Optimizing Functional Recellularization of Acellular Human Lung Scaffolds

优化非细胞人肺支架的功能再细胞化

• 批准号: 9250807
• 负责人: DANIEL J WEISS
• 金额: $ 38.13万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-15 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9250807
• 关键词: 3-Dimensional; AGTR2 gene; Address; Adult; Alveolar; Biological Assay; Biomedical Engineering; Blood Vessels; Cell Differentiation process; Cell physiology; Cells; Complex; Data; Detergents; Development; Dissection; Distal; Engineering; Environmental air flow; Epithelial; Epithelial Cells; Family suidae; Goals; Growth Factor; Human; Individual; Lung; Maintenance; Mechanical ventilation; Mechanics; Modeling; Perfusion; Periodicity; Play; Pleural; Primates; Protocols documentation; Publishing; Residual state; Rodent; Rodent Diseases; Rodent Model; Role; Scaffolding Protein; Scheme; Stem cells; Stretching; Structure of parenchyma of lung; Supporting Cell; Techniques; Transformed Cell Line; Vascular Endothelial Cell; base; cell growth; cell type; combinatorial; fetal; flexibility; induced pluripotent stem cell; innovation; novel; perfusion imaging; progenitor; public health relevance; scaffold; stem; surfactant production; tool

 DESCRIPTION: Despite exciting recent progress in ex vivo lung bioengineering utilizing decellularized whole lung scaffolds, a number of hurdles remain. These include schemes for functional recellularization with combinations of appropriate cell types, long term maintenance of recellularizing lungs, and incomplete understanding of optimal vascular perfusion and cyclic mechanical stretch influences on proliferation, differentiation, and appropriate function of cells inoculated into the scaffolds. Further, most progress to date has been made in rodent models. This in part reflects the relative difficulties in working with human as compared to rodent lungs including a more limited supply of human lungs and the practical aspects of handling larger more cumbersome lungs. This has hampered progress in assessing the multiple combinatorial conditions that must be evaluated for development of functional human lung tissue. To address these hurdles, we have developed novel and innovative techniques for studying recellularization of acellular human lung scaffolds. These include; a) More optimal detergent- based decellularization protocols; b) Infrared perfusion imaging and sophisticated mass spectrometric assessment of residual scaffold proteins; and c) Novel high throughput approaches for studying decellularized human lungs. This latter technique involves dissection of multiple small 1cm3 segments, each with a cannulatable bronchovascular bundle, from whole decellularized lungs. We have further developed a flexible artificial pleural coating for use with these segments that allows study of ventilation and perfusion of the recellularizing human lung segments. We have also developed a functional assay for surfactant production by assessing changes in lung mechanics in recellularizing scaffolds. Using these techniques, we have made significant advances in recellularizing acellular scaffolds produced from normal and diseased rodent, pig, primate, and human lungs. In particular, we have found significant effects of perfusion and of cyclic mechanical stretch on survival and differentiation, respectively, of pulmonary vascular endothelial cells and of type 2 alveolar epithelial cells in the scaffolds. The goal of the current application is to continue to develop advanced translational applicable schemes for recellularization of human lung scaffolds. Focus will be on combinations of relevant human lung cell types, including endogenous airway progenitor cells and iPS-derived lung epithelial cells (Specific Aim 1), developing appropriate perfusion schemes and perfusates for long term maintenance or recellularizing scaffolds (Specific Aim 2), and developing optimal mechanical ventilation schemes that will maximize influence of cyclic mechanical stretch on development of functional lung tissue (Specific Aim 3). Importantly, we have built an outstanding collaborative team with which to achieve these goals.

 描述：尽管使用脱细胞的整个肺部支架最近在体内肺生物工程中取得了令人兴奋的进展，但仍然存在许多障碍。这些方案包括用于功能性延迟的方案，结合了适当的细胞类型的组合，对肺部肺部的长期维持以及对最佳血管灌注和环状机械拉伸的不完全理解对接种到支架中的细胞的分化以及适当的细胞功能影响。此外，迄今为止，大多数进展是在啮齿动物模型中取得的。与啮齿动物肺部相比，这部分反映了与人合作的相对困难，包括人类肺的供应更有限，以及处理更大的繁琐肺部的实际方面。这阻碍了评估必须评估的多种组合条件以开发功能性人体肺组织。为了解决这些障碍，我们开发了用于研究细胞人类肺脚手架的新颖和创新技术。这些包括； a）更最佳的基于确定的脱细胞方案； b）残留支架蛋白的红外灌注成像和复杂的质谱评估； c）用于研究脱细胞的人肺的新型高通量方法。这项以后的技术涉及从整个脱细胞肺中解剖多个小1CM3段，每个段都有一个可固定的支气管束。我们进一步开发了一种柔性的人造胸膜涂层，用于这些片段，可以研究延迟人类肺部段的通风和灌注。我们还通过评估肺部脚手架中的肺力学变化来开发一种生存生产的功能评估。利用这些技术，我们在由正常和解散的啮齿动物，猪，灵长类动物和人类肺部产生的卵形冠状支架上取得了重大进步。特别是，我们发现灌注和环状机械拉伸对肺血管内皮细胞的生存和分化以及脚手架中2型肺泡上皮细胞的生存和分化的显着影响。电流的目标 应用程序将继续开发出高级翻译方案，以延迟人类肺部支架。 Focus will be on combinations of relevant human lung cell types, including endogenous airway progenitor cells and iPS-derived lung epithelial cells (Specific Aim 1), developing appropriate perfusion schemes and perfusates for long term maintenance or recellularizing scaffolds (Specific Aim 2), and developing optimal mechanical ventilation schemes that will maximize Influence of cyclic mechanical stretch on development of functional lung tissue (Specific Aim 3).重要的是，我们建立了一个杰出的合作团队，以实现这些目标。